Immunodeficient mice: the NOD connection

As we discussed in an earlier post, non-obese diabetic (NOD/ShiLtJ) mice are one of the most commonly used mice to model insulin-dependent (Type I) diabetes. It strikes me as strange that this strain would become the background of choice in the development of Nod Scid Gamma (NSG) and other severely immunodeficient mice that have revolutionized the development of next-generation human cancer and infectious disease models. How exactly did this happen?

Two distinct research goals drove the development of NOD immunodeficient models:

1) The development of a mouse that could be used to uncover which immune cells actually drive the development of autoimmune diabetes in NOD mice.

As outlined in a recent post, NOD-scid mice (NOD.CB17-Prkdcscid/J), because they are B and T cell-deficient, do not develop diabetes. As a result, they are the perfect hosts for the adoptive transfer of T and B cells from diabetic NOD mice to investigate their roles in the development of autoimmune (Type 1) diabetes.

2) The development of a mouse that would support long-term engraftment of human cells and tissues for studies related to human infectious disease and cancer.

The Prkdcscid is a spontaneous mutation initially characterized in a BALB/c congenic strain called C.B-17. Although C.B-17-scid mice are both T and B cell deficient, they fail to support long-term engraftment of the human immune cells that are critical for developing small animal models for human infectious diseases such as HIV. The likely explanation for their limited engraftment capacity is that C.B-17-scid mice retain normal NK and myeloid cell function.

It turns out that NOD inbred mice have inherent deficiencies in key humoral and cellular immune cell activities that enhances the overall immune deficiency of NOD-scid mice in comparison to C.B-17-scid or C57BL/6-scid (B6.CB17-Prkdcscid/SzJ) –mice.

NOD inbred mice are characterized by the following inherent immune deficiencies:

1) Absence of circulating complement. The complement system is a multiple protein-based component of the innate immune system that assists antibodies and phagocytic cells in destroying and clearing pathogens from the host.

Normally, a subgroup of complement proteins forms complexes that poke holes in a pathogen’s cellular membrane and lyse the invading cell. NOD mice, as well as several other inbred strains, are homozygous for a deletion in the hemolytic complement (Hc) gene that prevents Hc (aka C5) complement protein expression. Without functional C5, complement proteins fail to assemble, and invading pathogens escape their damaging effects.

Importantly, NK cell activity is a major impediment to the engraftment of human hematopoietic cells in mice. Therefore, the reduced NK cell function in NOD mice greatly improves engraftment of human cells.

3) Differentiation and functional deficits in macrophages and antigen presenting cells (APCs). Macrophages and APCs are immune cells that are important in engulfing pathogens. The subsequent displaying of antigens derived from the pathogens on the APC’s cell surface is key in triggering adaptive immune responses.

Combined, the above attributes make the NOD genetic background an ideal platform that, in combination with the scid mutation, produces mice that support better engraftment of human-derived immune cells. Indeed, NOD-scid mice support as much as 5-fold higher levels of human lymphoid cell engraftment, in comparison to for example C.B-17-scid mice.

Got Sirpa?

The race to develop even more advanced NOD-derived immunodeficient mice has been fueled by the need to develop “humanized” mice – that is, mice that express fully functional human immune cells in place of their murine counterparts.

It has been known for over a decade that immunodeficient mice with the NOD background engraft human hematopoietic stem cells (HSCs) more efficiently than immunodeficient mice on other genetic backgrounds. This phenomenon was explained recently by the discovery that macrophages in the bone marrow of NOD mice express a variant of the signal-regulatory protein alpha (Sirpa) that has a higher affinity for human hematopoietic stem cells (Takenaka et al. 2007; van den Berg and van der Schoot 2008). NOD-SCID mice that carry a Sirpa allele from NOR mice, a strain that is closely related to NOD, do not engraft hHSCs as readily.

What does Sirpa do?

SIRPA protein in both mouse and humans interacts with Cd47, and controls multiple functions in myeloid cells. One of the best documented is the negative regulation by SIRPA in macrophages of host cell phagocytosis, critical for “self-recognition” and transplantation tolerance. When macrophage-bound SIRPA binds to host cell Cd47, it generates a SHP-1-dependent inhibitory signal that prevents the macrophages from engulfing the Cd47-positive cell.

Presently, it is not clear, however, if the enhanced binding of SirpaNOD protein to hCD47 and the activation of a phagocytic inhibitory signal is the actual mechanism that is responsible for the enhanced engraftment of hHSCs in NOD-derived immunodeficient mice.

It is worth noting that immunodeficient mice on non-NOD-derived genetic backgrounds are now becoming available that overexpress the human SIRPA protein (e.g. C;129S4-Rag2tm1.1Flv Il2rgtm1.1FlvTg(SIRPA)1Flv/J) and show levels of hHSCs engraftement comparable to NSG mice.

The transfer of the scid mutation onto the NOD background proved to be a key breakthrough that both advanced our understanding of Type I diabetes and led to the development of more robust small animal models for studying a wide range of human diseases. Who knew that a diabetic mouse would become, arguably, the most important strain background in xenograft-based human disease research?